The Cytochalasins are fungal metabolites known to disassemble microfilamentous cell structures. We have found that cytochalasin D (CD) exerts an inhibitory effect on platelet functions which are mediated by contractile proteins. At a CD concentration of lug/ml of platelet- rich plasma (PRP), clot retraction was completely inhibited. Furthermore, at this concentration, the primary wave of platelet aggregation by ADP and Epinephrine was reduced, but the second wave was not affected. 14C-serotonin release was normal, however. Ultrastructurally, platelets were found to be highly vacuolated and devoid of granules. Nevertheless, they were metabolically active as evidenced by normal patterns of incorporation of (U14C) acetate and (UC14) d-glucose. In the presence of CD, the Ca2 plus ATPase activity of platelet myosin was inhibited, and the ability to crossreact with actin was also impaired. CD had no effect on the Mg2 ion and CA2 ion ATPase activity of platelet actomyosin, however. Because of these findings studies are now progressing to determine the participation of contractile proteins in platelet functions such as aggregation, secretion and clot retraction.